Cell Wall Surface Research Articles

Olpidiopsis sp., an oomycete from Madagascar that infects Bostrychia and other red algae: Host species susceptibility

SUMMARY Olpidiopsis sp. (Oomycota) was cultured with its original host Bostrychia moritziana (Sonder ex Kützing) J. Agardh from Madagascar. Bean‐shaped zoospores with two heterokont flagella attached to the host cell wall surface and in 2 days host cells began collapsing and one or more syncytia developed in each infected cell. Zoospores were cleaved and an exit tube with a small plug was formed. Complete development and zoospore discharge occurred in 3 days. Infection occurred in cells of polysiphonous branches, monosiphonous branches, rhizoids and reproductive stichidia. Dead cells of plants treated with microwave were not infected. Susceptibility was variable in other Bostrychia species from different countries. Bostrychia moritziana (Sonder ex Kützing) J. Agardh, and Bostrychia radicans (Montagne) Montagne from Madagascar were susceptible but one Bostrychia tenella (J. V. Lamouroux) J. Agardh isolate from Madagascar was susceptible and two were not. B. radicosa (Itono) J. A. West, G. C. Zuccarello et M. Hommersand isolates from Madagascar, Thailand, Australia and New Caledonia were susceptible but an isolate from Malaysia was not. B. radicans isolates from Mexico and Brazil were non‐susceptible as were Bostrychia flagellifera Post, Bostrychia harveyi Montagne, Bostrychia montagnei Harvey, Bostrychia simpliciuscula Harvey ex J. Agardh, Bostrychia tenuissima R. J. King et Puttock, Stictosiphonia intricata(Bory de Saint‐Vincent) P. C. Silva, Stictosiphonia kelanensis (Grunow) R. J. King et Puttock and Stictosiphonia tangatensis (Post) R. J. King et Puttock, Lophosiphonia sp., Neosiphonia sp. and Polysiphonia spp. isolates were also non‐susceptible. Many non‐susceptible strains showed initial cell‐collapse followed by rapid wound‐repair cell formation without syncytia or sporangia developing. Caloglossa leprieurii (Montagne) G. Martens from Madagascar showed cell‐collapse and wound‐repair in periaxial cells, but wing cells died and became purple without wound‐repair. Caloglossa ogasawaraensis Okamura and Caloglossa postiae M. Kamiya et R. J. King had no symptoms of infection. Dasysiphonia chejuensis I. K. Lee et J. A. West was not infected. Surprisingly, the conchocelis phase but not the blade phase of Porphyra pulchella J. A.West, G. C. Zuccarello and Porphyra suborbiculata Kjellman was infected. The conchocelis of Porphyra tenera Kjellman and Porphyra linearis Greville were infected but no blade stages were tested. Porphyra miniata (C. Agardh) C. Agardh and Porphyra dentata Kjellman conchocelis were not infected. Bangia atropurpurea (Roth) C. Agardh gametophyte filaments were not infected. Other red, brown and green algae were not infected. Time lapse videomicroscopy of development and spore release was done.

On the changes of pinewood (Pinus sylvestris L.) Chemical composition and ultrastructure during the attack by brown-rot fungi Postia placenta and Coniophora puteana

Abstract Pine sapwood blocks were exposed to brown-rot fungi Postia placenta and Coniophora puteana for 1, 2, 3, and 4 months. Ion exchange chromatography method was used to determine the changes in sugar content during brown-rot decay. The most remarkable feature was the preferred degradation of mannose both by P. placenta (80.9% weight loss) and C. puteana (77.5% weight loss) in comparison to the control. It can be interpreted as a result of the preferred degradation of the backbone chains of O -acetyl-galacto-glucomannans. For scanning electron microscopy, the wood degradation sequences obtained by the fracture method were used. The fractured surface of cell wall reflected the pattern of degradation of cellulose microfibrils. The surface of secondary wall decayed by P. placenta revealed both smooth and uneven regions throughout the test. C. puteana retained uneven cell wall surface only after the first month, while later the surface became smooth. This suggests that P. placenta tended to degrade cellulose amorphous regions more readily, while C. puteana , possessing full enzyme complement, was able to degrade both amorphous and crystalline regions more readily. The water vapor sorption method was applied for microstructural characteristics of wood. The formation of new pores of size 2.1–9.9 nm occurred during brown-rot decay, whose sizes and volume depend on the fungus culture and exposure time. It is assumed that the appearance of these pores is caused by the destruction of carbohydrates.

Lipid Headgroup Discrimination by Antimicrobial Peptide LL-37: Insight into Mechanism of Action

Interaction of the human antimicrobial peptide LL-37 with lipid monolayers has been investigated by a range of complementary techniques including pressure-area isotherms, insertion assay, epifluorescence microscopy, and synchrotron x-ray scattering, to analyze its mechanism of action. Lipid monolayers were formed at the air-liquid interface to mimic the surface of the bacterial cell wall and the outer leaflet of erythrocyte cell membrane by using phosphatidylglycerol (DPPG), phosphatidylcholine (DPPC), and phosphatidylethanolamine (DPPE) lipids. LL-37 is found to readily insert into DPPG monolayers, disrupting their structure and thus indicating bactericidal action. In contrast, DPPC and DPPE monolayers remained virtually unaffected by LL-37, demonstrating its nonhemolytic activity and lipid discrimination. Specular x-ray reflectivity data yielded considerable differences in layer thickness and electron-density profile after addition of the peptide to DPPG monolayers, but little change was seen after peptide injection when probing monolayers composed of DPPC and DPPE. Grazing incidence x-ray diffraction demonstrated significant peptide insertion and lateral packing order disruption of the DPPG monolayer by LL-37 insertion. Epifluorescence microscopy data support these findings.

Spiral structures of cotton fiber

In the present article, data from the microscopic investigation of the morphological structure of a cellular wall and plasmalemma of the Central Asian cotton fiber are given at different stages of their development. The objects under investigation were live hairs of five middle-fiber cotton species (Tashkent-1, 108-F, Namangan-77, Margelan-3, G. llirsutum L.) and the coarse-fiber variety of G. arboreum L., so-called Turfan Gtiza. To observe the cotton hair surface morphology, the original method of gelatinous prints was used. It is shown that, in all investigated samples of cotton cell-hairs, spiral structures are observed in the packing of cellulose microfibrils on the surface of the primary cell wall. Also, it is established that the plasmalemma of a cotton cell-hair makes a spiral movement along the direction of its growth. On the basis of the received data, a conclusion is made about the spiral mechanism for the deposition of cellulose microfibrils on a primary cell wall surface and the spiral movement of plasmalemma.

Genetic transformation of novel isolates of chicken Lactobacillusbearing probiotic features for expression of heterologous proteins: a tool to develop live oral vaccines

BackgroundThe use of lactic acid bacteria as vehicles to delivery antigens to immunize animals is a promising issue. When genetically modified, these bacteria can induce a specific local and systemic immune response against selected pathogens. Gastric acid and bile salts tolerance, production of antagonistic substances against pathogenic microorganisms, and adhesive ability to gut epithelium are other important characteristics that make these bacteria useful for oral immunization.ResultsBacteria isolated on de Man, Rogosa and Sharpe medium (MRS) from different gastrointestinal portions of broiler chicks were evaluated for their resistance to artificial gastric acid and bile salts, production of hydrogen peroxide, and cell surface hydrophobicity. Thirty-eight isolates were first typed at species level by PCR amplification of 16S-23S rRNA intergenic spacers using universal primers that anneal within 16S and 23S genes, followed by restriction digestion analyses of PCR amplicons (PCR-ARDRA). An expression cassette was assembled onto the pCR2.1-Topo vector by cloning the promoter, leader peptide, cell wall anchor and terminator sequences derived from the laminin binding S-layer protein gene of L. crispatus strain F5.7 (lbs gene). A sequence encoding the green fluorescent protein (GFP) was inserted as reporter gene, and an erythromycin resistance gene was added as selective marker. All constructs were able to express GFP in the cloning host E. coli XL1-Blue and different Lactobacillus strains as verified by FACS and laser scanning confocal microscopy.ConclusionLactobacillus isolated from gastrointestinal tract of broiler chickens and selected for probiotic characteristics can be genetically modified by introducing an expression cassette into the lbs locus. The transformed bacteria expressed on its cell wall surface different fluorescent proteins used as reporters of promoter function. It is possible then that similar bacterial model expressing pathogen antigens can be used as live oral vaccines to immunize broilers against infectious diseases.

The 24th International Specialized Symposium on Yeasts (ISSY 24) “Cell surface: a genomic and proteomic approach” Oropesa del Mar (Castellón, Spain), 28 September–2 October 2005

The 24th International Specialized Symposium on Yeasts was held under the auspices of the International Commission on Yeasts (ICY). Although initially focused on cell wall and cellular surfaces, the meeting was actually broader in scope and covered recent advances in the response to stress, signalling, and physiology of different fungi, which included both pathogenic and nonpathogenic yeasts as model systems for basic research and industrially oriented applications. The conference was dedicated to Prof. Jose Ruiz-Herrera , honouring his outstanding career and work on the biogenesis of the fungal cell wall. A broad range of experts in the field continued with their highly qualified contributions the tradition of the ISSY conferences to be an important convention for yeast scientists. We would like to highlight here some new data, ideas and trends that could be inferred from this meeting. Cell wall biogenesis attracted a considerable number of contributions, as this structure represents the interface between cells and their environment and is therefore per se a promising target for new therapeutic and diagnostic strategies. Elucidation of cell wall components and of the mechanisms leading to its construction is, as a consequence, a major objective in the field of yeast research. According to Miguel Gobernado (Hospital Santa Fe, Valencia, Spain), fungal infections nowadays are the third most frequent cause of hospital-acquired infections and, as the incidence of susceptible individuals may increase in time due to immunosuppressing medical treatments, its control is crucial. Antifungal therapy has relied on polyenes and azoles, but the first ones suffer from limited pharmacokinetics (partially overcome by the development of novel formulations like liposomal amphotericin B), while resistance to the azole family is emerging and may limit its usefulness in the future. For these reasons, the cell wall synthesis is an attractive area in which applied and basic research merge. …

Nesting Biologies and Immature Stages of the Tapinotaspidine Bee Genera Monoeca and Lanthanomelissa and of Their Osirine Cleptoparasites Protosiris and Parepeolus (Hymenoptera: Apidae: Apinae)

The nesting biologies of Monoeca haemorrhoidalis (Smith) and Lanthanomelissa betinae Urban (Tapinotaspidini) are described from southeastern Brazil. Both are ground nesting; the nests of the former are attacked by the cleptoparasite Protosiris gigas Melo (Osirini), and those of the latter are attacked by Parepeolus minutus Roig-Alsina (Osirini). Egg eclosion, larval feeding behavior, and cocoon spinning of M. haemorrhoidalis are detailed. A female of P. gigas opens the closed cell of M. haemorrhoidalis by making a large opening in the cell cap (which is plugged after ovipositioning) through which she apparently extends her metasoma. Indirect evidence suggests that she uses her metasomal apex, and perhaps even the sting, to kill the host egg or early instar. Protosiris eggs are either attached to the cell-wall surface of the nearly vertical host cells or dropped onto the surface of the provisions. First instars of P. gigas, with strongly curved, sharply pointed mandibles, are also capable of killing host immatures or competing cleptoparasites.Cocoons of all four species are compared and contrasted. The egg, all larval instars, and pupa of Monoeca haemorrhoidalis are described, as are the egg and postdefecating larva of Lanthanomelissa betinae. The egg, all larval instars, and pupa of Protosiris gigas are described, as is the postdefecating larva of Parepeolus minutus.Both Monoeca haemorrhoidalis and Protosiris gigas have four ovarioles per ovary. The egg indices and other ovarian features of both species are identified and discussed.The possible phylogenetic relationship of the Tapinotaspidini with the Osirini is briefly explored on the basis of data from this study. Possible phylogenetic relationships of the Osirini with other cleptoparasitic apids are analyzed.In the appendix,, the identity of the species of Monoeca, whose nesting biology is presented in the main paper, is discussed. The species is M. haemorrhoidalis (Smith, 1854), a species closely related to M. schrottkyi (Friese, 1902) and M. xanthopyga Harter-Marques, Cunha, and Moure, 2001. An identification key for distinguishing these three species is presented. Tetrapedia piliventris Friese is placed as a junior synonym of M. haemorrhoidalis (new synonymy). A lectotype is designated for Pachycentris schrottkyi Friese. The species of Protosiris found attacking M. haemorrhoidalis is here described as new, P. gigas Melo sp. nov. It is structurally most similar to P. caligneus (Shanks), from which it differs by its abundant yellow marks, plumose pubescence on the lower paraocular area, protruding anterior mesoscutum, and sparser punctation on the metasomal terga.Appendix: Taxonomic Notes on Monoeca and Description of a New Species of Protosiris, by Gabriel A. R. Melo

Removal of lead and copper ions from aqueous solutions by bacterial strain isolated from soil

Biosorption of Pb(II) and Cu(II) ions from aqueous solutions has been studied in a batch system by using a bacterial strain isolated from metal polluted soil. The bacterial strain was identified as Bacillus sp. The optimum conditions of biosorption were determined by investigating the initial pH, contact time and the initial concentrations of metal ions at constant temperature (25 ◦ C). The maximum biosorption of the metal ions was observed at pH 3.0 ± 0.1 for Pb(II) and pH 5.0 ± 0.1 for Cu(II) ions. Biosorption equilibrium times for Pb(II) and Cu(II) ions were observed in 15 and 30 min, respectively. The maximum biosorption capacities of Pb(II) and Cu(II) ions on Bacillus sp. were determined to be 92.27 ± 1.17 mg g −1 at 250 mg l −1 concentration and 16.25 ± 1.64 mg g −1 at 200 mg l −1 concentration, respectively. The experimental adsorption data were fitted to Langmuir isotherm model. Competition of metal ions during biosorption was investigated in binary metal solutions. The interactions between metal ions and functional groups on the cell wall surface of the biomass were confirmed by FTIR, SEM and EDAX analysis. The results indicated that bacterial isolate Bacillus sp. is a suitable biosorbent for the removal of Pb(II) and Cu(II) ions from aqueous solutions. © 2005 Published by Elsevier B.V.

How kaolinite plays an essential role in remediating oil-polluted seawater

AbstractAn investigation was carried out on the bioavailability of kaolinite and the role it plays in remediating oil-polluted seawater, since kaolinite is known to enable hydrocarbon-degrading bacteria to grow well. Experimental results revealed that significant amounts of Al and Si dissolved from kaolinite were not observed ((P > 0.05) in comparison with a control sample which contained no kaolinite) in the aqueous phase during the ~24 day experimental period. Transmission electron microscope observations and energy-dispersive spectroscope data revealed that some altered kaolinite particles appeared, connected to intact kaolinite and bacterial cells. Bacterial cells were associated and encrusted with intact and/or altered kaolinite clay particles, where mixed (C, O, Na and Si)-precipitates of kaolinite clays were formed on the surface of hydrocarbon-degrading bacterial cells. However, the uptake of Si (from kaolinite) by bacterial cells appeared to be more prevalent than Al, and there were no significant changes in basal spacings of kaolinite due to these altered kaolinite particles. Separate studies showed that hydrocarbon-degrading bacteria have a high resistance to Si. Thus, the present data suggest that Si from kaolinite may facilitate hydrocarbon-degrading bacterial growth as shown in our previous study (Chaerun et al., 2005), and the C-O-Na-Si complexes on the surfaces of bacterial cell walls may be the stimulator for hydrocarbon-degrading bacterial growth in seawater contaminated with oil spill.

Bioaccumulation of copper by Trichoderma viride

Studies were carried out on interaction of Trichoderma viride with copper and reports bioaccumulation as a mechanism of copper tolerance during growth. There was a marked increase in the lag phase of the growth, which was concentration dependent. At a concentration of 100 mg/L of CuCl 2 · 2H 2O, 81% of Cu(II) were removed by 3.4 g/L of the biomass in 72 h. The process was temperature and pH dependent. The maximum copper bioaccumulation occurred at 30 °C, pH 5.0. Metabolic inhibitors such as sodium azide (NaN 3) and 2,4-dinitrophenol (2,4-DNP) drastically reduced the extent of Cu(II) bioaccumulation. Electron microscopy and cell fractionation studies revealed that 70–80% of copper was present as a layer on the cell wall surface.

AFM study of crystalline cellulose in the cell walls of straw

AbstractCrystalline cellulose in the cell walls of straw was studied by atomic force microscopy (AFM). The samples were first treated and then observed by AFM under dimethylsulfoxide (DMSO). The crystalline regions were located and two allomorphs of crystalline cellulose, triclinic Iα and monoclinic Iβ phases, were identified. In most crystalline regions, the Iα and Iβ phases are intimately associated, with the Iβ phase more abundant than the Iα phase. In some small domains only one phase with long‐range order was observed. It was demonstrated that in these one‐phase domains, Iα phase crystals always have their (010) plane lying parallel to the cell wall surface and Iβ phase crystals with (110) plane lying parallel. Copyright © 2005 Society of Chemical Industry

Characterization of surface sugars on algal cells with fluorescein isothiocyanate-conjugated lectins

We used qualitative and quantitative fluorescence microscopy of the fluorescein isothiocyanate-conjugated lectins Concanavalin A, phytohaemagglutinin-erythroagglutinin, pokeweed mitogen, and peanut agglutinin to examine sugar composition on the cell surface and cell-associated mucilage (where present) in a number of cultured and environmental algae. Lectin-binding activity was markedly different between laboratory-cultured and environmental samples of the same species. Sugar composition of the cyanobacterium Anabaena cylindrica varied with growth cycle, although no clear pattern of change was observed. Akinetes typically showed lectin-binding activity higher than that of the vegetative cells or heterocysts throughout the growth cycle. Algae with mucilage showed greater lectin binding, indicating that mucilage contained more surface sugars accessible to the lectin probe compared with the cell wall surface. A low level of galactose and N-acetyl galactosamine (detected by peanut agglutinin) was associated with the surface mucilage of most algal species. Relatively high amounts of mannose, glucose, and N-acetyl glucosamine (detected by Concanavalin A, phytohaemagglutinin, and pokeweed mitogen) were also present. Lectin binding was shown to be a highly specific and sensitive approach to the examination of cell surface chemistry of both cultured and environmental algae and to the study of biodiversity in phytoplankton.

Arabinogalactan proteins are required for apical cell extension in the moss Physcomitrella patens.

Cell biological, structural, and genetic approaches have demonstrated the presence of arabinogalactan proteins (AGPs) in the moss Physcomitrella patens and provided evidence for their function in cell expansion and specifically in the extension of apical tip-growing cells. Inhibitor studies indicated that apical cell expansion in P. patens is blocked by synthetic AGP binding beta-glucosyl Yariv reagent (betaGlcYR). The anti-(1-->5)-alpha-L-arabinan monoclonal antibody LM6 binds to some AGPs in P. patens, to all plasma membranes, and to the cell wall surface at the most apical region of growing protonemal filaments. Moreover, LM6 labeling of cell walls at the tips of apical cells of P. patens was abolished in the presence of betaGlcYR, suggesting that the localized movement of AGPs from the plasma membrane to the cell wall is a component of the mechanism of tip growth. Biochemical and bioinformatic analyses were used to identify seven P. patens ESTs encoding putative AGP core proteins from homology with Arabidopsis thaliana, Brassica napus, and Oryza sativa sequences and from peptide fragments isolated from betaGlcYR-precipitated AGPs. Gene knockout by homologous recombination of one of these genes, P. patens AGP1, encoding a classical AGP core protein, resulted in reduced cell lengths in protonemal filaments, indicating a role for AGP1 in apical cell expansion in P. patens.

Monoclonal Immunoglobulin G1 Directed againstAspergillus fumigatusCell Wall Glycoprotein Protects against Experimental Murine Aspergillosis

Most of the biological functions related to pathogenicity and virulence reside in the fungal cell wall, which, being the outermost part of the cell, mediates the host-fungus interplay. For these reasons much effort has focused on the discovery of useful inhibitors of cell wall glucan, chitin, and mannoprotein biosynthesis. In the absence of a wide-spectrum, safe, and potent antifungal agent, a new strategy for antifungal therapy is directed towards the development of monoclonal antibodies (MAbs). In the present study the MAb A9 (immunoglobulin G1 [IgG1]) was identified from hybridomas raised in BALB/c mice immunized with cell wall antigen of Aspergillus fumigatus. The immunoreactive epitopes for this IgG1 MAb appeared to be associated with a peptide moiety, and indirect immunofluorescence microscopy revealed its binding to the cell wall surface of hyphae as well as with swollen conidia. MAb A9 inhibited hyphal development as observed by MTT [3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay (25.76%), reduced the duration of spore germination, and exerted an in vitro cidal effect against Aspergillus fumigatus. The in vivo protective efficacy of MAb A9 was also evaluated in a murine model of invasive aspergillosis, where a reduction in CFU (>4 log(10) units) was observed in kidney tissue of BALB/c mice challenged with A. fumigatus (2 x 10(5) CFU/ml) and where enhanced mean survival times (19.5 days) compared to the control (7.1 days) and an irrelevant MAb (6.1 days) were also observed.

Effects of media composition on submerged culture spores of the entomopathogenic fungus, Metarhizium anisopliae var. acridum, Part 1: Comparison of cell wall characteristics and drying stability among three spore types

Metarhizium anisopliae var. acridum (IMI 330189) can produce at least three spore types in vitro; blastospores, submerged conidia, and aerial conidia, as defined by culturing conditions, sporogenesis, and spore morphology. This study compares morphological characteristics (dimensions and cell wall structure), chemical properties of cell wall surfaces (charge, hydrophobicity, and lectin binding), and performance (germination rate and drying stability) among these three spore types. Submerged conidia and aerial conidia both possessed thick, double-layered cell walls, with hydrophobic regions on their surfaces. However, in contrast to aerial conidia, submerged conidia have: (1) a greater affinity for the lectin concanavalin-A; (2) more anionic net surface charge; and (3) a less distinct outer rodlet layer. Blastospores were longer and more variable in length than both submerged conidia and aerial conidia, and had thinner single-layered cell walls that lacked an outer rodlet layer. Also, blastospores had a greater affinity than either conidia type for the lectin, wheat germ agglutinin. Blastaspores lacked hydrophobic regions on their surface, and had a lower anionic net surface charge than submerged conidia. In culture, blastospores germinated the fastest followed by submerged conidia, and then aerial conidia. Survival of submerged conidia and aerial conidia were similar after drying on silica gel, and was greater than that for blastospores. We provide corroborating information for differentiating spore types previously based on method of production, sporogenesis, and appearance of spores. These physical characteristics may have practical application for predicting spore-performance characteristics relevant to production and efficacy of mycoinsecticides.

Effects of media composition on submerged culture spores of the entomopathogenic fungus, Metarhizium anisopliae var. acridum Part 2: Effects of media osmolality on cell wall characteristics, carbohydrate concentrations, drying stability, and pathogenicity

This study evaluates osmolality of a submerged conidia-producing medium in relation to the following spore characteristics: yield, morphology (dimensions and cell wall structure), chemical properties of cell wall surfaces (charge, hydrophobicity, and lectin binding), cytoplasmic polyols and trehalose, and performance (drying stability and pathogenicity). Spore production was increased by the addition of up to 150 g l−1 polyethylene glycol 200 (PEG). Spores from high osmolality medium (HOM spores) containing 100 g l−1 PEG had thin cell walls and dimensions more similar to blastospores than submerged conidia or aerial conidia. However, a faint electron-dense layer separating primary and secondary HOM spores’ cell walls was discernable by transmission electron microscopy as found in aerial and submerged conidia but not found in blastospores. HOM spores also appeared to have an outer rodlet layer, unlike blastospores, although it was thinner than those observed in submerged conidia. HOM spores’ surfaces possessed hydrophobic microsites, which was further evidence of the presence of a rodlet layer. In addition, HOM spores had concentrations of exposed N-acetyl-β-d-glucosaminyl residues intermediate between blastospores and submerged conidia potentially indicating a masking of underlying cell wall by a rodlet layer. All spore types had exposed α-d-mannosyl and/or α-d-glucosyl residues, but lacked oligosaccharides. Similar to blastospores, HOM spores were less anionic than submerged conida. Although HOM spores had thin cell walls, they were more stable to drying than blastospores and submerged conidia. Relative drying stability did not appear to be the result of differences in polyol or trehalose concentrations, since trehalose concentrations were lower in HOM spores than submerged conidia and polyol concentrations were similar between the two spore types. HOM spores had faster germination rates than submerged conidia, similar to blastospores, and they were more pathogenic to Schistocerca americana than submerged conidia and aerial conidia.

Oxygen-releasing step of ZnFe 2O 4/(ZnO + Fe 3O 4)-system in air using concentrated solar energy for solar hydrogen production

The oxygen-releasing step of the ZnFe 2O 4/(ZnO + Fe 3O 4)-system for solar hydrogen production with two-step water splitting using concentrated solar energy was studied under the air-flow condition by irradiation with concentrated Xe lamp beams from a solar simulator. The spinel-type compound of ZnFe 2O 4 (Zn-ferrite) releases O 2 gas under the air-flow condition at 1800 K and then decomposes into Fe 3O 4 ( = Fe II Fe 2 III O 4 ) and ZnO with a nearly 100% yield (ZnFe 2O 4 = ZnO + 2/3Fe 3O 4 + 1/6O 2). The ZnO was deposited as the thin layer on the surface of the reaction cell wall. A thermodynamic study showed that the ZnO was produced by the reaction between the O 2 gas in the air and the metal Zn vapor generated from ZnFe 2O 4. With the combined process of the present study on the O 2-releasing step and the previous one on the H 2 generation step (ZnO + 2/3Fe 3O 4 + 1/3H 2O = ZnFe 2O 4 + 1/3H 2) for the ZnFe 2O 4/(ZnO + Fe 3O 4)-system, solar H 2 production was demonstrated by one cycle of the ZnFe 2O 4/(ZnO + Fe 3O 4)-system, where the O 2-releasing step had been carried out in air at 1800 K and the H 2 generation step at 1100 K.

Hydrous ferric oxide precipitation in the presence of nonmetabolizing bacteria: Constraints on the mechanism of a biotic effect

We have used room temperature and cryogenic 57Fe Mössbauer spectroscopy, powder X-ray diffraction (pXRD), mineral magnetometry, and transmission electron microscopy (TEM), to study the synthetic precipitation of hydrous ferric oxides (HFOs) prepared either in the absence (abiotic, a-HFO) or presence (biotic, b-HFO) of nonmetabolizing bacterial cells ( Bacillus subtilis or Bacillus licheniformis, ∼10 8 cells/mL) and under otherwise identical chemical conditions, starting from Fe(II) (10 −2, 10 −3, or 10 −4 mol/L) under open oxic conditions and at different pH (6–9). We have also performed the first Mössbauer spectroscopy measurements of bacterial cell wall ( Bacillus subtilis) surface complexed Fe, where Fe(III) (10 −3.5–10 −4.5 mol/L) was added to a fixed concentration of cells (∼10 8 cells/mL) under open oxic conditions and at various pH (2.5–4.3). We find that non-metabolic bacterial cell wall surface complexation of Fe is not passive in that it affects Fe speciation in at least two ways: (1) it can reduce Fe(III) to sorbed-Fe 2+ by a proposed steric and charge transfer effect and (2) it stabilizes Fe(II) as sorbed-Fe 2+ against ambient oxidation. The cell wall sorption of Fe occurs in a manner that is not compatible with incorporation into the HFO structure (different coordination environment and stabilization of the ferrous state) and the cell wall-sorbed Fe is not chemically bonded to the HFO particle when they coexist (the sorbed Fe is not magnetically polarized by the HFO particle in its magnetically ordered state). This invalidates the concept that sorption is the first step in a heterogeneous nucleation of HFO onto bacterial cell walls. Both the a-HFOs and the b-HFOs are predominantly varieties of ferrihydrite (Fh), often containing admixtures of nanophase lepidocrocite (nLp), yet they show significant abiotic/biotic differences: Biotic Fh has less intraparticle (including surface region) atomic order (Mössbauer quadrupole splitting), smaller primary particle size (magnetometry blocking temperature), weaker Fe to particle bond strength (Mössbauer center shift), and no six-line Fh (6L-Fh) admixture (pXRD, magnetometry). Contrary to current belief, we find that 6L-Fh appears to be precipitated directly, under a-HFO conditions, from either Fe(II) or Fe(III), and depending on Fe concentration and pH, whereas the presence of bacteria disables all such 6L-Fh precipitation and produces two-line Fh (2L-Fh)-like biotic coprecipitates. Given the nature of the differences between a-HFO and b-HFO and their synthesis condition dependences, several biotic precipitation mechanisms (template effect, near-cell environment effect, catalyzed nucleation and/or growth effect, and substrate-based coprecipitation) are ruled out. The prevailing present view of a template or heterogeneous nucleation barrier reduction effect, in particular, is shown not to be the cause of the large observed biotic effects on the resulting HFOs. The only proposed mechanism (relevant to Fh) that is consistent with all our observations is coprecipitation with and possible surface poisoning by ancillary bacteriagenic compounds. That bacterial cell wall functional groups are redox active and the characteristics of biotic (i.e., natural) HFOs compared to those of abiotic (i.e., synthetic) HFOs have several possible biogeochemical implications regarding Fe cycling, in the photic zones of water columns in particular.

Transmission Electron Microscopy Study of Enterohemorrhagic Escherichia coli O157:H7 in Apple Tissue

We investigated the ability of enterohemorrhagic Escherichia coli O157:H7 to spread in wounded apple tissue by transmission electron microscopy. Red Delicious apples were wounded with an artist knife (7 mm depth) and either inoculated with 10μl per wound of decimally diluted E. coli O157:H7 or submerged into E. coli O157:H7 suspended in sterile distilled water and then stored at 37°C for 24 h. Transmission electron microscopy showed E. coli O157:H7 formed bacterial aggregates near the apple cell walls, and single cells were in close proximity to the apple cell wall surfaces and to plasma membranes. E. coli O157:H7 presence caused degradation of plasma membranes and release of the cytoplasm contents of the apple cortical cells into the central vacuole. Apple tissue turgor pressure tests showed that the apple cells infected with E. coli O157:H7 isolates were more likely to rupture than the control noninoculated apple cells. E. coli O157:H7 cells grown in apple tissue showed the formation of granules and vesicles within the bacterial cytoplasma and separation of the plasma membranes. Our study shows that E. coli O157:H7 can grow and survive in the apple tissue environment by causing degradation of the apple cellular components.

Cell surface display system for Lactococcus lactis: a novel development for oral vaccine

The food-grade Lactococcus lactis is a potential vector to be used as a live vehicle for the delivery of heterologous proteins for vaccine and pharmaceutical purposes. We constructed a plasmid vector pSVac that harbors a 255-bp single-repeat sequence of the cell wall-binding protein region of the AcmA protein. The recombinant plasmid was transformed into Escherichia coli and expression of the gene fragment was driven by the T7 promoter of the plasmid. SDS-PAGE showed the presence of the putative AcmA' fragment and this was confirmed by Western blot analysis. The protein was isolated and purified using a His-tag affinity column. When mixed with a culture of L. lactis MG1363, ELISA and immunofluorescence assays showed that the cell wall-binding fragment was anchored onto the outer surface of the bacteria. This indicated that the AcmA' repeat unit retained the active site for binding onto the cell wall surface of the L. lactis cells. Stability assays showed that the fusion proteins (AcmA/A1, AcmA/A3) were stably docked onto the surface for at least 5 days. The AcmA' fragment was also shown to be able to strongly bind onto the cell surface of naturally occurring lactococcal strains and Lactobacillus and, with less strength, the cell surface of Bacillus sphericus. The new system designed for cell surface display of recombinant proteins on L. lactis was evaluated for the expression and display of A1 and A3 regions of the VP1 protein of enterovirus 71 (EV71). The A1 and A3 regions of the VP1 protein of EV71 were cloned upstream to the cell wall-binding domains of AcmA protein and successfully expressed as AcmA/A1 and AcmA/A3. Whole-cell ELISA showed the successful display of VP1 protein epitopes of EV71 on the surface of L. lactis. The success of the anchoring system developed in this study for docking the A1 and A3 epitopes of VP1 onto the surface of L. lactis cells opens up the possibilities of peptide and protein display for not only Lactococcus but also for other gram-positive bacteria. This novel way of displaying epitopes on the cell surface of L. lactis and other related organisms should be very useful in the delivery of vaccines and other useful proteins.